Cytoplasmic islet cell antibodies recognize distinct islet antigens in IDDM but not in stiff man syndrome

Cytoplasmic islet cell antibodies are well-established predictive markers of IDDM. Although target molecules of ICA have been suggested to be gangliosides, human monoclonal ICA of the immunoglobulin G class (MICA 1-6) produced from a patient with newly diagnosed IDDM recognized glutamate decarboxylase as a target antigen. Here we analyzed the possible heterogeneity of target antigens of ICA by subtracting the GAD-specific ICA staining from total ICA staining of sera. This was achieved 1) by preabsorption of ICA+ sera with recombinant GAD65 and/or GAD67 expressed in a baculovirus system and 2) by ICA analysis of sera on mouse pancreas, as GAD antibodies do not stain mouse islets in the immunofluorescence test. We show that 24 of 25 sera from newly diagnosed patients with IDDM recognize islet antigens besides GAD. In contrast, GAD was the only islet antigen recognized by ICA from 7 sera from patients with stiff man syndrome. Two of these sera, however, recognized antigens besides GAD in Purkinje cells. In patients with IDDM, non-GAD ICA were diverse. One group, found in 64% of the sera, stained human and mouse islets, whereas the other group of non-GAD ICA was human specific. Therefore, mouse islets distinguish two groups of non-GAD ICA and lack additional target epitopes of ICA besides GAD. Longitudinal analysis of 6 sera from nondiabetic ICA+ individuals revealed that mouse-reactive ICA may appear closer to clinical onset of IDDM in some individuals

Determination of islet cell antibodies using an ELISA system with a preparation of rat insulinoma (RIN A2) cells

An enzyme-linked immunosorbent assay (ELISA) was established for the detection of islet cell antibodies in human sera. The antigen was prepared from rat insulinoma (RIN A2) cells. Cells were dissociated in lysis buffer and the lysate was centrifuged at 100,000 x g. The supernatant was used to coat microtiter ELISA plates (10 micrograms protein/ml in PBS pH 7.2). Non-specific binding sites on the plates were blocked with 2% PBS-BSA. Human test sera were preabsorbed on separate plates using 2% PBS-BSA and incubated on precoated plates at an optimal dilution of 1/10 in 60 mM PBS for 60 min at 37 degrees C. Phosphatase-labeled anti-human IgG serum and phosphatase substrate were applied and the reaction was stopped by adding 3 M NaOH. Out of 90 sera from type I diabetic patients, 47 (52.2%) reacted in the new ELISA whereas none of 15 type II diabetics, 50 sera containing non-islet specific antibodies or 100 normal controls were positive. In the same group of patients, ICA were positive in 63.3%. When both, the ELISA and conventional ICA testing were applied, the number of positives was increased to 83%. The ICA-ELISA with the above described antigen preparation provides a well standardized and reproducible test method which is highly specific for type I diabetes. It may therefore be useful for large screening procedures

Ground-motion prediction equations for southern Spain and southern Norway obtained using the composite model perspective

In this paper, two sets of earthquake ground-motion relations to estimate peak ground and response spectral acceleration are developed for sites in southern Spain and in southern Norway using a recently published composite approach. For this purpose seven empirical ground-motion relations developed from recorded strong-motion data from different parts of the world were employed. The different relations were first adjusted based on a number of transformations to convert the differing choices of independent parameters to a single one. After these transformations, which include the scatter introduced, were performed, the equations were modified to account for differences between the host and the target regions using the stochastic method to compute the host-to-target conversion factors. Finally functions were fitted to the derived ground-motion estimates to obtain sets of seven individual equations for use in probabilistic seismic hazard assessment for southern Spain and southern Norway. The relations are compared with local ones published for the two regions. The composite methodology calls for the setting up of independent logic trees for the median values and for the sigma values, in order to properly separate epistemic and aleatory uncertainties after the corrections and the conversions

Violence as an Under-Recognized Barrier to Women’s Realization of Their Right to Adequate Food and Nutrition: Case Studies From Georgia and South Africa

This article addresses under-acknowledged barriers of structural violence and discrimination that interfere with women’s capacity to realize their human rights generally, and their right to adequate food and nutrition in particular. Case studies from Georgia and South Africa illustrate the need for a human rights–based approach to food and nutrition security that prioritizes non-discrimination, public participation, and self-determination. These principles are frustrated by different types of structural violence that, if not seriously addressed, pose multiple barriers to women’s economic, public, and social engagement. </jats:p

Baculovirus-Mediated Expression of Human 65 kDa and 67 kDa Glutamic Acid Decarboxylases in SF9 Insect Cells and Their Relevance in Diagnosis of Insulin-Dependent Diabetes Mellitus

cDNAs coding for the full-length human 65 and 67 kDa glutamic acid decarboxylases (GAD65 and GAD67) were amplified from pancreas and hippocampus cDNA libraries by polymerase chain reaction, respectively. Both cDNAs were inserted into a baculovirus vector which mediated highly efficient expression of the human GAD65 and GAD67 with histidine-hexapeptides as affinity ligands at their C-termini in Spodoptera frugiperda (Sf9) cells. The recombinant GAD proteins were purified to homogeneity by affinity chromatography using a metal-chelating matrix. The infected Sf9 insect cells expressed the recombinant human GAD65 and GAD67 with natural-like conformations, as confirmed by measurement of their enzyme activities as well as their fully restored autoantigenicities. Immunoprecipitation of metabolically labeled infected Sf9 cells demonstrated the autoantigenic potential of the recombinant GAD proteins. The practicability of using recombinant GAD65 and GAD67 derived from the baculovirus expression system for the development of an immunoassay for the diagnosis of insulin-dependent diabetes mellitus is discussed

Epidemiology and immunogenetic background of islet cell antibody - positive nondiabetic schoolchildren : Ulm-Frankfurt population study

Islet cell antibodies (ICAs) were determined in a large cohort of white nondiabetic schoolchildren (n = 4287) from a homogenous population in southern Germany. The prevalence of ICA levels greater than or equal to 5 Juvenile Diabetes Foundation (JDF) U was 1.05% (95% confidence interval 0.8-1.4%). Analysis of HLA-DR beta and -DQ beta alleles revealed that the specificities found to be increased in insulin-dependent (type I) diabetic subjects with the same ethnic background were also associated with ICA positivity in the nondiabetic schoolchildren. HLA-DR3 (P less than 0.01) and -DR4 (P less than 0.01) phenotypes and absence of Asp residue (P less than 0.01) at codon 57 of the HLA-DQ beta-chain were significantly increased in ICA+ compared with control subjects. High levels of ICAs, which were categorized as either greater than or equal to 17 or greater than or equal to 30 JDF U, were found to be associated with amino acids other than Asp at position 57 of the HLA-DQ beta-chain. No association of ICA level was found for HLA-DR phenotypes

Application of Single-Station Sigma and Site-Response Characterization in a Probabilistic Seismic-Hazard Analysis for a New Nuclear Site

Aleatory variability in ground-motion prediction, represented by the standard deviation (sigma) of a ground-motion prediction equation, exerts a very strong influence on the results of probabilistic seismic-hazard analysis (PSHA). This is especially so at the low annual exceedance frequencies considered for nuclear facilities; in these cases, even small reductions in sigma can have a marked effect on the hazard estimates. Proper separation and quantification of aleatory variability and epistemic uncertainty can lead to defensible reductions in sigma. One such approach is the single-station sigma concept, which removes that part of sigma corresponding to repeatable site-specific effects. However, the site-to-site component must then be constrained by site-specific measurements or else modeled as epistemic uncertainty and incorporated into the modeling of site effects. The practical application of the single-station sigma concept, including the characterization of the dynamic properties of the site and the incorporation of site-response effects into the hazard calculations, is illustrated for a PSHA conducted at a rock site under consideration for the potential construction of a nuclear power plant.Civil, Architectural, and Environmental Engineerin